key: cord-0779337-stpcj8x1
authors: Patel, Manvendra; Chaubey, Abhishek Kumar; Pittman, Charles U.; Mlsna, Todd; Mohan, Dinesh
title: Coronavirus (SARS-CoV-2) in the Environment: Occurrence, Persistence, Analysis in Aquatic Systems and Possible Management
date: 2020-10-02
journal: Sci Total Environ
DOI: 10.1016/j.scitotenv.2020.142698
sha: d2168c7e8be8e4340ea3b9dd48a11b3461d8df38
doc_id: 779337
cord_uid: stpcj8x1

The year 2020 brought the news of the emergence of a new respiratory disease (COVID-19) from Wuhan, China. The disease is now a global pandemic and is caused by a virus named SARS-CoV-2 by international bodies. Important viral transmission sources include human contact, respiratory droplets and aerosols, and through contact with contaminated objects. However, viral shedding in feces and urine by COVID-19-afflicted patients raises concerns about SARS-CoV-2 entering aquatic systems. Recently, targeted SARS-CoV-2 genome fragments have been successfully detected in wastewater, sewage sludge and river waters around the world. Wastewater-based epidemiology (WBE) studies can provide early detection and assessment of COVID-19 transmission and the growth of active cases within given wastewater catchment areas. WEB surveillance's ability to detect the growth of cases was demonstrated. Was this science applied throughout the world as this pandemic spread throughout the globe? Wastewater treatment efficacy for SARS-CoV-2 removal and risk assessments associated with treated water are reported. Disinfection strategies using chemical disinfectants, heat and radiation for deactivating and destroying SARS-CoV-2 are explained. Analytical methods of SARS-CoV-2 detection are covered. This review provides a more complete overview of the present status of SARS-CoV-2 and its consequences in aquatic systems. So far, WBE programs have not yet served to provide the early alerts to authorities that they have the potential to achieve. This would be desirable in order to activate broad public health measures at earlier stages of local and regional stages of transmission.

With the end of 2019, a respiratory coronavirus disease (COVID-19) outbreak emerged from Wuhan city in China (WHO, 2020a) . This illness is caused by a new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (WHO, 2020a). The outbreak of COVID-19 has become a universal pandemic since March 11, 2020 (WHO, 2020a . By 18 September 2020, a total of 30,055,710 affected patients were confirmed and 943,433 deaths were reported due to COVID-19 illness worldwide (WHO, 2020b), although both these numbers are surely much higher, especially the former.

Starting from Wuhan in December 2019, the virus had spread to 216 countries by the date this paper was submitted (WHO, 2020b) . Respiratory droplets along with direct contacts (touching an infected person or contacting contaminated objects and then transmitting the virus from one"s hand to the mouth, nose and eyes are main SARS-CoV-2 transmission routes (Chan et al., 2020b; Meselson, 2020) . Recent evidence shows the presence of SARS-CoV-2 in human urine and feces Tang et al., 2020; Wölfel et al., 2020; Zhang et al., 2020b) . Human shedding of the virus through nose, mouth, urine and feces led to the presence of SARS-CoV-2 in the environment. SARS-CoV-2 has been reported in air, on household objects including door knobs, taps, and handles (Cai et al., 2020b; Chan et al., 2020b; Chin et al., 2020) .

CoV-2published papers available as per "web of science" database, (B) SARS-CoV-2 preprints on medRxiv and bioRxiv database, (C) peer reviewed "SARS-CoV-2" articles (data available as per different mentioned keys) on "web of science" database, (D) "SARS-CoV-2" preprints (data available as per different mentioned keys) on medRxiv and bioRxiv database,

To date a large number of viewpoints, short communications and reviews have been published on SARS-CoV-2 and coronaviruses in the environment. Reviews include topics such as occurrence in water and wastewater sources (Foladori et al., 2020; Núñez-Delgado, 2020) , transmission (Heller et al., 2020; Hindson, Hata and Honda, 2020; Michael-Kordatou et al., 2020; Naddeo and Liu, 2020; Sims and Kasprzyk-Hordern, 2020) and disinfection strategies (Chauhan, 2020; Kamp, 2020; Kampf et al., 2020b) . None of these reviews compile all the issues to provide a complete picture of the issue of SARS-CoV-2 in the environment, with a special focus on water and wastewater. This review provides a more complete overview of SARS-CoV-2 occurrence, persistence, analysis and disinfection in water and wastewater sources. Data compiled and presented in many of these prior reviews are on postulated and derived from the studies of other coronaviruses including SARS and MERS. In contrast, this review work provides SARS-CoV-2 information available in literature prior to the manuscript submission.

Web of Science and Google scholar were used for peer reviewed literature selection.

Literature from bioRvix and medRvix were used for preprint collection for writing this review. According to Web of Science, bioRxiv and medRxiv data, more than 16,500 preprints and peer reviewed articles have been published on SARS-CoV-2 up to September, 2020. Only 62 peer reviewed papers and 500 preprints were focused on the keyword "SARS-CoV-2 in water" and "SARS-CoV-2 in wastewater" combined. A large number of preprints were not included in the study as these are not directly related to the topic of this review. Approximately 150 articles, preprints and reports were selected based on their relevance to SARS-CoV-2 in water and wastewater. A total of 60 peer reviewed articles/reviews together with 50 relevant preprints published in 2020 were cited. In addition, 45 other relevant articles published before 2020 were also discussed.

J o u r n a l P r e -p r o o f Journal Pre-proof

Severe respiratory syndrome coronavirus 2 (SARS-CoV-2) is a member of the Coronaviridae family and the Nidovirales order and the sub family Coronavirinae (Harapan et al., 2020) . This virus belongs to genera Betacoronavirus which also includes SARS-CoV-1, HCoV-OC43, MERS-CoV and HCoV-HKU1 (Harapan et al., 2020) . SARS-CoV-2 is responsible for COVID-19. Coronaviruses are enveloped single stranded RNA viruses (size 60-220 nms) with crown like structures on their surface (Naddeo and Liu, 2020; Rosa et al., 2012) . Transmission routes involve human-tohuman spread that occurs mainly through aerosol droplets from mouth and nose of the infected person. High viral loads have been found in the respiratory tract of infected individuals.

Therefore, viral transmission from patient to surrounding air and onto objects is an important route of transmission. The median time of SARS-CoV-2 detection in feces (22 days) is higher than in serum samples (16 days) and in respiratory airways (18 days) in COVID-19 patients . Another study reports persistent SARS-CoV-2 shedding in patient"s feces for up to 33 days after being tested negative for respiratory viral RNA (Wu et al., 2020b) . The large viral load in urine and feces leads to SARS-CoV-2 presence in terrestrial and aquatic sources in the environment (Ahmed et al., 2020a; Arora et al., 2020; Kocamemi et al., 2020a; Kocamemi et al., 2020b; Medema et al., 2020) . According to another study, SARS-CoV-2 can remain active for up to 25 days in water sources (Shutler et al., 2020) . This study also estimates that contaminated water sources (water systems, waterways and rivers) can deliver the equivalent of >100 SARS-CoV-2 copies with 100 mL or less water in the countries with J o u r n a l P r e -p r o o f high SARS-CoV 2 prevalence (Shutler et al., 2020) . Asymptomatic persons can also disseminate coronaviruses through second-hand aerosols (SHA) and second-hand smoke (SHS) from cigarettes and combustible tobacco products (Mahabee-Gittens et al., 2020) . Therefore, more comprehensive assessments of occurrence, persistence, its analysis and management strategies are necessary.

In general, viral persistence in a given environment is essential for its transmission. However, the environmental presence of a virus is dependent upon several factors. SARS-CoV-2 has been detected on surfaces including cell phones, door handles and many other day-to-day items (Aboubakr et al., 2020) Only limited data are available until now regarding the SARS-CoV-2 persistence on various materials in the environment (Table 1) . Extensive literature search points toward a deficiency of available data concerning SARS-CoV-2 persistence in aquatic systems.

Stability in air and on surfaces is a prominent factor determining the efficiency of SARS-CoV-2 transmission (Doremalen et al., 2020) . Contaminated dry surfaces also play an important role in SARS-CoV-2 transmission (Kamp, 2020) . With the enormous number of 10 8 viral copies in just one mL of sputum, SARS-CoV-2 can rapidly infect numerous people (Rothe et al., 2020) . Different coronaviruses have been reported to persist between 2h to 9 days on different surfaces (Kampf et al., 2020a) . The SARS-CoV-2 half-life in aerosol and on copper, cardboard, polypropylene and stainless steel are 2. 74, 3.4, 8.45, 15.9 and 13.1 h, respectively (Doremalen et al., 2020) . SARS-CoV-2 persistence in two aerosols and on several different surfaces is summarized in Table 1. J o u r n a l P r e -p r o o f SARS-CoV-2 persisted for 3h in aerosol at 21-23 °C and 65% relative humidity (Doremalen et al., 2020) . Another study asserted that the SARS-CoV-2"s prevalence and aerosol stability in ambient environmental conditions (23 °C and ~53 % relative humidity and in the absence of UV) lasted >16 hours (Fears et al., 2020) . Infectious SARS-CoV-2 was detected for the entire 16 h period, and a minor but constant SARS-CoV-2 fraction maintained its replication-competence (Fears et al., 2020) . Thus, this virus can be considered an airborne pathogen for entire 16 hours. These authors also assessed the qualitative integrity of SARS-CoV-2 after longer-term aerosol experimentation through scanning electron microscopy (SEM) imaging (Fears et al., 2020) . SARS-CoV-2 is heterogeneous and ovoid in shape. It maintained its shape, size and morphologies for the entire 16 hours, which is consistent with the aerosol suspension stability experiment (Fears et al., 2020) . The wide variations in data of these two available studies demonstrate the lack of reliability and proper statistical analysis in the available data. However, both studies concluded that SARS-CoV-2 aerosols can remain pathogenic for several hours.

Significant contamination of common household objects including remote control, mobile phones, bed table, toilets, washbasins, bed rails, window ledges, ventilation grates and floor under patient"s bed by SARS-CoV-2 is reported (Cai et al., 2020a; Ong et al., 2020; Santarpia et al., 2020) . SARS-CoV-2 contamination at room sites (87%), toilet sites (such as sink, toilet bowl and door handle) (60%) was reported for a mild symptomatic patient (Ong et al., 2020) . Transmission of SARS-CoV-2 to humans from different contaminated surfaces is also reported (Aboubakr et al., 2020; Cahill and Morris, 2020; Núñez-Delgado, 2020) , thus evaluation of viral persistence on everyday J o u r n a l P r e -p r o o f encountered common surfaces becomes essential. SARS-CoV-2 survival on some important surfaces is provided in Table 1 . Plastic surfaces are common objects, often discarded in water in our society (Geyer et al., 2017; Hale and Song, 2020) . Plastic bags and other plastic substances can carry bacteria and viruses with them (Hale and Song, 2020) . Reusable bags and other common plastic-based products are easily contaminated and have the potential to spread coronaviruses, since SARS-CoV-2 can survive between 4-7 days (Chin et al., 2020; Doremalen et al., 2020) . This is another reason they need to be kept out waterways. Proper disposal of single use plastics including PPE, gloves, gowns, syringes used by patients and their caretakers in medical facilities is necessary. SARS-CoV-2 was reported to survive for more than 7 days on layers of surgical masks (Chin et al., 2020) . Table 1 documents SARS-CoV-2 survival studies on other household objects including paper (3 h), tissue paper (3 h), copper (8 h), cardboard (2 d), cloth (2 d), wood (2 d), banknote paper (4 d) and stainless steel (4 d). However, these studies do not mimic the real environmental situations by using higher (>10 4 ) numbers of infectious virus particles in a small study area (Goldman, 2020) . Therefore, the chances of depositing highly infectious viral particle concentrations (>10 4 ) on fomite surfaces are minimal thus, the chances of viral transmission from fomites are low (Goldman, 2020) .

Regardless of SARS-CoV-2 survival on various surfaces, low transmission possibilities through fomites exists (Goldman, 2020) . Till date no clear evidences are available for infectious potential of SARS-CoV-2 from different surfaces (Cai et al., 2020a; Cai et al., 2020b) . However, Middle East respiratory syndrome (MERS) coronavirus can remain infectious up to 60 minutes after aerosolization (Cai et al., 2020a) . This point J o u r n a l P r e -p r o o f substantiate the possible infectious potential of SARS-COV-2 from fomites (Cai et al., 2020a) . A study conducted in a shopping mall in Wenzhou, China, demonstrated the SARS-CoV-2 spread through fomites e.g. restroom taps or elevator buttons (Cai et al., 2020a) . This study indicated low intensity SARS-CoV-2 transmission by indirect conveyance routes such as fomites (Cai et al., 2020a) .

The persistence of various coronaviruses have been reported in both treated and untreated water (Carraturo et al., 2020; Gundy et al., 2009) . The persistence of SARS-CoV-2 in aquatic systems is largely unknown and undocumented. Survival and sustainability of SARS-CoV-2 in aqueous systems is influenced by initial viral load, type of medium, temperature, organic matter, presence of biologic fluids and with the presence of organic and inorganic substances (Carraturo et al., 2020; Romano-Bertrand et al., 2020) . Since coronaviruses are highly sensitive to temperature, changes can cause drastic survival time differences (Naddeo and Liu, 2020) . Coronavirus (HCoV 229E), a SARS-CoV-2 surrogate, can survive (99.9 % inactivation) up to 588 d at 4 °C in filtered tap water (Gundy et al., 2009 ). However, this survival level (99.9% inactivation) reduced to just 10.1 d at 23 °C (Gundy et al., 2009) . Similarly, survival times of HCoV 229E in unfiltered tap water, filtered primary effluent, unfiltered primary effluent and secondary effluents at 23°C are 12.1, 2.35, 3.54, and 2.77 days, respectively (Gundy et al., 2009) . Although, SARS-CoV-2 inactivation studies are still lacking, based on other coronaviruses studies similar persistences are expected (Carraturo et al., 2020; Gundy et al., 2009) . SARS-CoV-2 persistence in a viral transport medium (concentration 6.8 log TCID 50 /mL) is exponentially reduced with a rise in temperature (Chin et al., 2020) . Only 0.7 log TCID 50 /mL (10 % approx.) reduction in J o u r n a l P r e -p r o o f SARS-CoV-2 concentration was achieved at 4 °C in 14 days, which was reduced to 14 days at 22 °C (Chin et al., 2020) . Further, SARS-CoV-2 survivability progressively dropped to 2 days, 30 minutes and 5 minutes for 37, 56 and 70 °C, respectively (Chin et al., 2020) .

Exposure to sunlight or UV light drastically limits coronavirus survival, as is the case for many microorganisms (Naddeo and Liu, 2020) . Shielding viruses from light exposure and viral settling behavior (settling of virus in aquatic sources with time, as well as with suspended load) are both enhanced by the presence of organic matter.

SARS-CoV-2 can attach itself to organic matter particles and settle quite easily (Naddeo and Liu, 2020) . The presence of antagonist microorganisms can decrease the viral survival (Naddeo and Liu, 2020) . Coronavirus studies suggested extremely low SARS-CoV-2 survival occurred especially in wastewater temperatures of > 20 °C (Collivignarelli et al., 2020) . In contrast to that report, viable SARS-CoV-2 has also been reported in Indian wastewaters during the peak of summer (with ambient temperatures as high as 40-45 °C) (Arora et al., 2020; Kumar et al., 2020) . This raises concerns. A variety of further studies are needed on the stability and survival of SARS-CoV-2 in aqueous systems.

Recently, a computational model was developed to estimate the SARS-CoV-2 persistence in wastewater (Hart and Halden, 2020a) . This model estimates the half-life of SARS-COV-2 by considering an exponential viral decay dependent on wastewater temperature (Hart and Halden, 2020a; Hart and Halden, 2020b; Hart and Halden, 2020c) . Based on this model the SARS-CoV-2 half-life is estimated between 4.8 and 7.2 J o u r n a l P r e -p r o o f Journal Pre-proof h at 20 °C. The 99.9% reduction time ranges between 48 and 72 h at 20 °C for SARS-CoV-2 in wastewater (Hart and Halden, 2020a) . 

Viral shedding in urine and feces is likely the biggest source of viral RNA in water and wastewater systems. An estimated load of 0.056 to 11.3 billion SARS-CoV-2 genomes/infected person/per day is injected into wastewater (Hart and Halden, 2020a) .

Coronavirus RNA shedding has earlier been reported for SARS-CoV and MERS-CoV through feces (Corman et al., 2016; Leung et al., 2003) . As many as 10 7 and 2.5 × 10 4 SARS-CoV RNA copies/mL were reported in case of diarrhea and urine respectively J o u r n a l P r e -p r o o f Journal Pre-proof (Hung et al., 2004) . Persistent SARS-CoV-2 RNA shedding has been also reported in 27-89 % of COVID-19 patients" excreta specimens including anal/rectal swabs and feces (Cai et al., 2020b; Holshue et al., 2020; Tang et al., 2020; Wölfel et al., 2020; Zhang et al., 2020b) . As many as 10 8 viral RNA copies per gram of feces were reported in several studies (Lescure et al., 2020; Pan et al., 2020; Wölfel et al., 2020) . SARS-CoV-2 RNA fecal shedding can last upto seven weeks after onset of first symptoms has also been reported in clinical studies (Cai et al., 2020b; Wu et al., 2020b; Xiao et al., 2020) . Viral RNA was shed through feces by 81.8% of cases even after patients received negative results from throat swab tests (Ling et al., 2020) . Furthermore, the feces of asymptomatic patients were also found positive for SARS-CoV-2 RNA (Mizumoto et al., 2020; Nishiura et al., 2020; Tang et al., 2020) .

Coronaviruses were detected previously in sewage, water and wastewater sources (Hung et al., 2004; Leung et al., 2003) as already mentioned. SARS-CoV-2 enters these waters from human urine and feces. The fecal-oral route can be important for SARS-CoV-2 transmission and future investigations on the possibilities of SARS-CoV-2 fecal-oral transmission should incorporate environmental studies to ascertain the possible conditions favoring such transmission (Yeo et al., 2020) . At present none of the available studies have provided proof of fecal-oral transmission of SARS-CoV-2 (Cahill and Morris, 2020; Foladori et al., 2020; Gupta et al., 2020; Hindson, 2020; Sehmi and Cheruiyot, 2020) . However, the presence of viable SARS-CoV-2 in wastewater and water sources point toward a potential transmission of the virus through contaminated aerosols (Foladori et al., 2020; Gupta et al., 2020; Heller et al., 2020; Hindson, 2020; Sehmi and Cheruiyot, 2020; Shutler et al., 2020) . Three possible primary pathways J o u r n a l P r e -p r o o f were proposed for fecal-oral SARS-CoV-2 transmissions (Heller et al., 2020) . The first is direct contact with contaminated water; the second is through vectors including insects;

the third is through surfaces which came in contact with contaminated water or surfaces contaminated by vectors (Heller et al., 2020) .

SARS coronaviruses are reported to remain infectious for up to 4 days in stool samples (Weber et al., 2016) . Of particular significance, coronaviruses can also remain active and infectious in sewage and water for several days and weeks (Casanova et al., 2009; Gundy et al., 2009) . Similarly, high percentages (up to 99%) coronaviruses can remain viable for several days in tap waters and in sewage effluents at room temperature (Casanova et al., 2009; Gundy et al., 2009) . No studies have performed specific SARS-CoV-2 systematic survival time determinations in water and wastewater systems up to the date of the current review. SARS-CoV-2 can remain active for a long as 14 and 2 days at 22ºC and 37ºC, respectively, in viral transport medium (Chin et al., 2020) . On the basis of these viral transport results, SARS-CoV-2 significant survival times in water and wastewater systems are thought to be likely (Shutler et al., 2020) .

That study also concludes that SARS-CoV-2 survival is temperature driven and decreases with increase in temperature. SARS-CoV-2 can contribute to transmission above detection levels and remains active for as long as 25 days at 5 °C in wastewater (Shutler et al., 2020) .

Recent studies also suggested possible SARS-CoV-2 cross-transmissions occurred between 9 patients when bath center in Huai"an, Jiangsu province, China (Table 2) . These studies have also found correlations between the number of COVID-19 cases and the amount of SARS-CoV-2 RNA fragments present in wastewater (Medema et al., 2020) . Furthermore, SARS-CoV-2 RNAs were also found in the household wastewaters of home quarantined COVID-19 affected persons (Döhla et al., 2020) . These wastewater sample sources include washbasin siphons, shower siphons and toilets (Döhla et al., 2020) . While these findings allow one method of mapping COVID-19 cases in a location, the key question is: what do these detected SARS-CoV-2 RNAs mean for human transmission in each location they are found? What virus loads were present?

Wide variations in intact SARS-CoV-2 detection frequencies and its loads in wastewater have been observed (Table 2) . Higher viral concentrations were expected and reported from the countries with high COVID-19 caseloads, such as France, Japan,

Turkey and USA (Table 2) . Similarly, locations with high COVID-19 cases also show higher detection frequencies for SARS-CoV-2 RNA in Brazil (Fongaro et al., 2020) ,

France (Foladori et al., 2020; Rimoldi et al., 2020) , India (Arora et al., 2020; Kumar et al., 2020) , Turkey (Kocamemi et al., 2020a) and USA (Sherchan et al., 2020) (Table 2 ).

E.g. during February and March, only 29 COVID-19 cases were reported in Milan.

SARS-CoV-2 presence in wastewaters also shows a similar pattern with a sample occurrence frequency of 2/8 (25%) and this was only reported on February 24 th and 28 th (La Rosa et al., 2020) . However, when high numbers of COVID-19 cases occurred in Rome between March 31 st and April 2 nd , the SARS-CoV-2 sample detection frequency was 4/4 (100 %) (Rosa et al., 2020b) . However, high numbers of COVID-19 cases in Rome, SARS-CoV-2 detection frequency translates to 4/4 (100 %) samples between

March 31 st and April 2 nd (Rosa et al., 2020b) . Similar results were also reported in other studies from Japan (Haramoto et al., 2020) , Netherlands (Medema et al., 2020) , Brazil (Fongaro et al., 2020) , India (Arora et al., 2020) . Thus, wastewater analysis can provide accurate predictions of epidemiological cases in a regional level.

The SARS-CoV-2 presence in river waters was also examined and reported (Guerrero-Latorre et al., 2020; Haramoto et al., 2020) . River water samples in Yamanshi prefecture of Japan tested negative for the presence of SARS-CoV-2 (Haramoto et al., 2020) . The low COVID-19 prevalence in the studied region was the suggested explanation (Haramoto et al., 2020) . However, in high COVID-19 prevalence regions, as in case of the urban rivers of Quito (Ecuador), all river water samples were tested positive for SARS-CoV-2 (Guerrero-Latorre et al., 2020). Both N1 and N2 target genome assays were utilized for SARS-CoV 2 RNA evaluations in urban river waters.

Urban river water in Quito contained 2.84 x 10 5 to 3.19 x 10 6 N1 and 2.07 x 10 5 to 2.23

x 10 6 for N2 target genome copies/Liter (Guerrero-Latorre et al., 2020) . This study provided correlations with COVID-19 active cases 14 days prior the sampling study (Guerrero-Latorre et al., 2020) .

Like wastewater, sewage sludge is also known to host a wide variety of human viruses as well as recent strains of circulating coronavirus (Bibby and Peccia, 2013) . infections as in asymptomatic and presymptomatic cases. SARS-CoV-2 monitoring in sludge would be advantageous, as sludge is a concentrated and well-mixed sample (Peccia et al., 2020) . Therefore, analyzing sewage for SARS-CoV-2 can become an early indicator for outbreak dynamics assessment within a community (Peccia et al., 2020) . Primary sludge samples were collected daily from a wastewater treatment facility serving nearly 200,000 residents from March 19, 2020 to May 1, 2020 during a Covid-19 outbreak area of metropolitan area of New Haven, Connecticut (CT), USA (Peccia et al., 2020) . All the tested samples show the existence of SARS-CoV-2 RNA (values vary from 1.7 x 10 3 to 4.6 x 10 5 copies per mL). Results were also quantitatively compared with new COVID-19 cases and community hospital admission data. SARS-CoV-2 RNA amounts in sludge exhibits a high correlation (R 2 > 0.97) with new COVID-19 cases and hospital admissions data (Peccia et al., 2020) . The SARS-CoV-2 RNA presence in primary sludge is 2-3 times higher than in wastewater with no treatment (Peccia et al., 2020) . Therefore, sludge can archive the SARS-CoV-2 presence with COVID-19

infections with a high correlation (Michael-Kordatou et al., 2020; Peccia et al., 2020) .

J o u r n a l P r e -p r o o f sampling were carried out (Nemudryi et al., 2020) . Composite samples provide more reliable data for SARS-CoV-2 analysis (Nemudryi et al., 2020) . Large variations in sample collection volume (36 ml to 2 L) have been reported (Table 3) 

Sample preparation/pre-conditioning is an important step before sample concentration can be done. Pre-conditioning steps enhances viral recoveries and overall concentration efficiencies. Common pre-conditioning steps include pre-filtration and sample pH and salinity adjustment are reported elsewhere (Bofill-Mas and Rusiñol, 2020). MgCl 2 , beef extract, and glycine are commonly used agents of pre-conditioning to enhance viral elution (Bofill-Mas and Rusiñol, 2020).

Common sample concentration approaches prior to quantification have been recently reviewed (Bofill-Mas and Rusiñol, 2020; Haramoto et al., 2020) . Several virus concentration methods with specific advantages and disadvantages can be found in J o u r n a l P r e -p r o o f Table 4 . Common methods includes precipitation/flocculation (using an organic flocculant or ammonium sulfate precipitation), adsorption-elution (negatively/positively charged filters, glass powder or fiber), ultracentrifugation, ultrafiltration, lyophilisation and filtration (Bofill-Mas and Rusiñol, 2020; Bosch et al., 2006) .

Surrogate viruses are often used when studying uncultivable viruses (Bosch et al., 2006) . The murine hepatitis virus, for example, is commonly used for persistence and recovery studies as human corona virus surrogate due to their structural similarities (Ahmed et al., 2020b; Casanova et al., 2009; Patel et al., 2017; Ye et al., 2016) .

Another advantage is the non-pathogenic nature of these surrogate viruses towards humans which also reduces the need for the highest levels of biosafety precautions (Ahmed et al., 2020b) . Adsorption-extraction methods with MgCl 2 pre-treatment (65.7 ± 23.8%) and without any pretreatment (60.5 ± 20.2%) were the most efficient concentration methods (Ahmed et al., 2020b) .

Centrifugation or filtration to remove debris, electronegative membrane filtration (Peccia et al., 2020) or virus is eluted by PEG precipitation from the matrix are applied (Balboa et al., 2020) . Organic compounds (such as humic substances) can interfere with downstream/in vitro viral detection via co-concentration with viral RNA . Advantages and disadvantages of various concentration procedures are provided in Table 4 .

Sample volume is another important factor affecting virus detection results. In general, enteric viruses detection in untreated wastewater samples normally used <100 mL samples for concentration (Haramoto et al., 2018) . J o u r n a l P r e -p r o o f

Polymerase chain reactions (PCR) based techniques including quantitative PCR (qPCR) and reverse transcription quantitative PCR (RT-qPCR) are widely used for RNA and DNA viruses quantification in wastewater (Farkas et al., 2020a; Haramoto et al., 2018) . These methods are useful for small viral genome segment detection. PCR based methods provide rapid, accurate and sensitive strain-level detections for up to five targets in one assay (Jiang et al., 2014) . Several designed RT-qPCR assays were applied for SARS-CoV-2 detection (Chan et al., 2020a; Nalla et al., 2020; Vogels et al., 2020) , which also provide satisfactory results in wastewater monitoring (Ahmed et al., 2020a; Medema et al., 2020; Wurtzer et al., 2020a; Wurtzer et al., 2020b) . A SARS-CoV-2 RNA detection study conducted in Australian wastewaters used the N_Sarbeco assay (Ahmed et al., 2020a) , CDC-N1, -N2, -N3 and the E_Sarbeco assays was used in a Dutch study (Medema et al., 2020) and CDC-N1, -N2 and -N3 assays in a Spanish study (Randazzo et al., 2020a; Randazzo et al., 2020b) . Different SARS-CoV-2 genome sections used in these assays are provided in Table 2 . However, dissimilar assays may provide varying performance for viral detection. Use of different primer/probes for quantification showed substantial differences in rates of viral detection. For example, the "N1" and "N3" genes were detected for SARS-CoV-2 analysis in wastewater (positive) while the "N2" assay did not (Medema et al., 2020) . Hence, multiple primer/probe sets are recommended for usage. The presence of organic cocontaminants limits the use of qPCR-based methods by inhibiting polymerase enzymes and reverse transcription .

Digital PCR (d-PCR) were also reported for the estimation of viruses in environmental samples (Farkas et al., 2020a) . This method provides absolute target quantification and minimizes inhibition. However, digital PCR is more costly than quantitative PCR analysis. Biosensors and isothermal amplification for detection and quantification of viral DNA/RNA in environmental samples are also emerging techniques. These techniques provides results within an hour (Farkas et al., 2020a) . Paper-based microfluidics devices are another easy and inexpensive platform with the potential to rapidly detect viruses in wastewaters (Mao et al., 2020) . Antibody based detection techniques involving immunoassay techniques including colloidal gold immunoassays, enzyme linked immunosorbent assays (ELISA), lateral flows immunoassays, time-resolved fluorescence immunoassays are also in development along with a variety of other antibody based detection kits available in market (Jalandraad et al., 2020 (Jalandraad et al., 2020) . However, the low sensitivity of these assays versus traditional PCR-based methods is a disadvantage. These methods were also not rigorously tested in the field (Ishii et al., 2014; Jalandraad et al., 2020) .

Thus, RT-qPCR based techniques provide the most reliable results to date.

Quality control and quality assurance (QA/QC) is important for analytical methods. Collected samples must be true representatives and proper precautions are necessary for pre-conditioning and concentration steps. Method concentration J o u r n a l P r e -p r o o f efficiencies need to be determined properly before applications with real world samples.

Only methods with high recoveries should to be applied for wastewater analysis.

Method repeatability and reproducibility must be assessed properly. Artificially contaminated samples are often used to evaluate detection methods and the use of surrogate viruses is a well explored practice. Surrogates are very helpful when addressing problems with uncultivable (not easily cultivable) viruses (Bosch et al., 2006) . Special precaution are needed in wastewater samples because the presence of turbidity and organic matter can affect sample concentration (Bofill-Mas and Rusiñol, 2020).

The development of standard methods is required to accurately evaluate viral concentration for accurate evaluation of WBE and environmental surveillance. At present no widely accepted standard method/procedure exists for wastewater SARS-CoV-2 detection (Collivignarelli et al., 2020) , however, modified standard methods have been employed. For example, WHO"s standard poliovirus surveillance procedure (WHO, 2003) , was modified to develop a standardized method for SARS-CoV-2 RNA analysis in Italian wastewaters (Rosa et al., 2020b) . The development of similar standardized methods for wastewater SARS-CoV-2 RNA detection could fulfill significant analytical research gaps for wastewater surveillance. Standard methods should also be able to quantify viruses in complex wastewater matrixes and thus would provide results that could be compared from samples collected around the world (Collivignarelli et al., 2020) .

Identifying symptomatic COVID-19 infected individuals is straightforward, while identification of presymptomatic and asymptomatic individuals is a bigger concern. The median COVID-19 incubation period with no symptoms is 5.1 days (Lauer et al., 2020) .

Between 18-31% percent of infected patients are reported to be asymptomatic (Mizumoto et al., 2020; Nishiura et al., 2020) . Asymptomatic individuals also shed SARS-CoV-2 virions in their feces . These presymptomatic and asymptomatic patients are a major source of untraced COVID-19 transmissions (Hata and Honda, 2020) . The lack of proper instrumentation and high costs limits diagnostic testing in many countries. This problem is more serious in developing countries. Thus, limited diagnostic testing coupled with the presence of presymptomatic and asymptomatic patients has led to uncertainties of the extent COVID-19 spread in many regions (Bivins et al., 2020) .

Water and wastewater systems harbor numerous pathogenic microorganisms (Adriaenssens et al., 2018) . Enveloped viruses, including coronaviruses, inactivate rapidly in water and wastewater without a host when compared to other viruses (Casanova et al., 2009; Casanova et al., 2010; Rosa et al., 2020a) . Nevertheless, they were present in many wastewaters due to the continuous SARS-CoV-2 influx by humans through urine and excreta (Sehmi and Cheruiyot, 2020; Singer and Wray, 2020; Sun et al., 2020) (Table 2) . SARS-CoV-2 contamination of water supplies have the potential to infect whole communities . A super-spreading event previously reported for a SARS spread in Hong Kong was related to faculty sewage system Gormley et al., 2012; J o u r n a l P r e -p r o o f al., 2014; Peiris et al., 2003; WHO, 2003) . About 342 SARS cases were reported from high rise building in Amory Garden, Hong Kong. Epidemiological studies suggested the role of faulty sewage system contaminated with the excreta on "index patient" in causing this super spreading event Peiris et al., 2003) .This faulty sewage system facilitated transmission of virus-laden droplets through aerosols as well as contaminating surfaces in the bathrooms (Peiris et al., 2003; WHO, 2003) . Aerosols derived from wastewaters through leakage, flushing and malfunctioning sewer plumbing facilities are identified as possible infection route during SARS spread in 2003 (D. Nghiem et al., 2020) . Norovirus transmission through wastewater flow and wastewater derived aerosols was also reported (Gormley et al., 2014) . Toilet flushing can generate aerosols with airborne pathogens including E.coli, Staphylococcus epidermidis, Pseudomonas alcaligenes as well as viruses (Lai et al., 2018; Li et al., 2020; Wang et al., 2020a) . A recent study also reported the SARS-CoV-2 spread through wastewater plumbing systems in Guangzhou, China Kang et al., 2020) .

The ubiquitous SARS-CoV-2 presence in human excreta offers the potential of using viral RNA sewage surveys to estimate the epidemiological status of COVID-19"s prevalence in a region (Bivins et al., 2020) . This epidemiological monitoring is known as wastewater-based epidemiology (WBE) or environmental surveillance (Bivins et al., 2020; Daughton, 2020a; Daughton, 2020b; Sims and Kasprzyk-Hordern, 2020) .

Effective screening of suspected infectious individuals from every individual household remains a tough, highly challenging logistical task for medical professionals.

It is highly labor intensive, time-consuming and costly. WBE could be an alternative and J o u r n a l P r e -p r o o f effective method for SARS-CoV-2 local and regional assessments (Daughton, 2020a; Daughton, 2020b; Hata and Honda, 2020) . Temporal changes of viral concentrations in wastewater using WBE can provide information related to a specific viral absence or presence, outbreak dynamics and its demographic and human health effects . A 2013 example of silent transmissions and wild poliovirus type 1 reintroductions were observed in Israel during routine sewage samples surveillance, without any clinically reported case appearing (Manor et al., 2014) . In another example, the first SARS-CoV-2 case was reported in Italy on February 21 st , 2020 and the first report of SARS-CoV-2 being present in wastewater was reported shortly after, on

February 24 th (Rosa et al., 2020b) .

This wastewater report also suggested that SARS-CoV-2 infections might have started before the detection of the first case in Italy (Rosa et al., 2020b) . COVID-19

cases have been correlated with the SARS-COV-2 presence in wastewater from various WWTP catchment areas by several studies (Ahmed et al., 2020a; Haramoto et al., 2020; Medema et al., 2020; Randazzo et al., 2020a; Randazzo et al., 2020b; Rosa et al., 2020b; Wurtzer et al., 2020b) . These studies have successfully demonstrated the wastewater surveillance"s potential to provide epidemiological dynamics assessments to better understand and design outbreak handling approaches. For example, SARS-CoV-2 analysis within sewage treatment plants of the Valencian metropolitan area (~1,200,000 inhabitants) in Spain, provided a direct correlation with declared active cases in the region (Randazzo et al., 2020a) . This study illustrated that SARS-COV-2 detection in wastewaters coincided with the emergence of the first case in the region.

The rise in COVID-19 cases also correlates with the rise in SARS-CoV-2 RNA"s presence in wastewater (Randazzo et al., 2020a) .

Regions with only a few COVID-19 cases have also reported SARS-CoV-2 load in wastewaters (Haramoto et al., 2020; Randazzo et al., 2020b; Wurtzer et al., 2020b) .

Wastewater surveillance also provided the proof of SARS-CoV-2 RNA detection weeks before the actual confirmation of cases in the region of Murcia (Spain) had occurred (Randazzo et al., 2020b) . and costly due to sample collection and testing compared to environmental surveillance, and d) environmental surveillance collects information about symptomatic, presymptomatic and asymptomatic patients, which clinical surveillance cannot provide (Hata and Honda, 2020) . Even a single infected individual can be detected within a population of 2,000,000 individuals (Hart and Halden, 2020a) . Globally, 2.1 billion people can be monitored with the existing 105,600 WWTPs (Hart and Halden, 2020a ).

Thus, WBE should become an essential tool for global pandemic/epidemic analysis and prevention.

As established above, SARS-CoV-2 contaminated water and wastewaters have the potential to spread COVID-19. Thus, assessing SARS-CoV-2 removal using existing techniques is important. These include secondary and tertiary treatments in WWTPs and advanced drinking water treatments. The use of disinfecting agents also needs to be evaluated. Heat and radiation are also important methods for microbial removal and viral removal from aqueous systems (Arora et al., 2020; Chin et al., 2020) . Use of high temperature on SARS-CoV-2 inactivation and rapid SARS-CoV-2 removal using bleach, benzyl alkyl ammonium chloride, chloroxylenol, ethanol and povidine-iodine are well known (Chin et al., 2020) . Many disinfectants, including hypochlorites, quaternary ammonium salts, hydrogen peroxide, peracetic acids, mono persulfates, and chlorine dioxide are listed by United States Environmental Protection Agency for SARS-CoV-2 J o u r n a l P r e -p r o o f disinfection (USEPA, 2020). Successful SARS-CoV-2 inactivation in viral transport medium was achieved using common disinfectants including household bleach (both 1:49 and 1:99 ratios in water), benzylkonium chloride (0.1 %), chloroxylenol (0.05 %), chlorhexidine (0.05 %), ethanol (70 %) and povidone-iodine (7.5 %) within 5 minutes (Chin et al., 2020) . However, only 54 % inactivation can be achieved with hand soap solution (1:49) in 5 minutes and 100 % inactivation is observed in 15 minutes (Chin et al., 2020) . Thus, common disinfectants can also be applied for wastewater disinfection.

For example, sodium hypochlorite was successfully applied for large scale SARS-CoV-2 loaded hospital wastewater treatment in Wuhan, China (Zhang et al., 2020a) . Potential of other techniques have also been suggested .

Coronaviruses have low survival rates in high temperature waters (Gundy et al., 2009 (Gundy et al., 2009) . Preliminary studies also consider low SARS-CoV-2 RNA survival times in wastewaters above 20 °C (Collivignarelli et al., 2020) . SARS-CoV-2 added into both raw and treated wastewater samples 48 and 72 hours after inoculation displayed insignificant vitality and no cytopathic effects on Vero E6 cells (Rimoldi et al., 2020) .

Several studies revealed SARS-CoV-2 RNA detection in wastewaters recently, but few of these have investigated SARS-CoV-2 RNA removal from wastewater in treatment plants (Arora et al., 2020; Collivignarelli et al., 2020; Kumar et al., 2020; Randazzo et al., 2020a; Randazzo et al., 2020b; Zhang et al., 2020a) . However, some J o u r n a l P r e -p r o o f studies evaluated the presence of SARS-CoV-2 RNA in both influents and effluents of WWTPs (Arora et al., 2020; Collivignarelli et al., 2020; Kumar et al., 2020; Randazzo et al., 2020a; Randazzo et al., 2020b; Zhang et al., 2020a) . Table 5 summarizes SARS-CoV-2 RNA concentrations in both influent and effluent wastewaters versus the WWTPs" treatment techniques. These studies indicated declines in SARS-CoV-2 RNA in WWTP effluents compared to the corresponding influents (Table 5 ). For example, 42

influents, 18 secondary and 12 tertiary treated effluents were collected and analyzed for SARS-CoV-2 (Randazzo et al., 2020b) . Of these, only 35 of 42 (85 %) influent samples tested SARS-CoV-2 positive compared to only 2 of 18 (11 %) of the secondary-treated samples (Randazzo et al., 2020a ). This provides insights into possible SARS-CoV-2 removal within WWTPs. (Zhang et al., 2020a) UASB-Up-flow anaerobic sludge blanket, MBBR-Moving bed biofilm reactor, SBR-Sequencing batch Reactor J o u r n a l P r e -p r o o f

The efficacy of Up-flow anaerobic sludge blanket (UASB) and aeration pond-based secondary treatments to remove SARS-CoV-2 RNA was demonstrated by influent and effluent sample concentration analyses conducted on 08/05/2020 and 27/05/2020 in Pirana, Ahmedabad in Gujarat, India (Kumar et al., 2020) . All influent samples tested SARS-CoV-2 positive with a maximum concentration of 2.419 × 10 8 genome copies/L (Kumar et al., 2020) . However, all effluent samples tested negative for SARS-CoV-2 RNA (Kumar et al., 2020) . A study employing a moving bed biofilm reactor (MBBR) and a sequencing batch reactor (SBR) treatment also turned SARS-CoV-2 positive untreated wastewater samples into a SARS-CoV-2 negative effluents (Arora et al., 2020) .

Application of treated wastewater for agricultural purposes and in gardening might have public health risks, when contaminated with SARS-CoV-2 (Arora et al., 2020) . Similarly, application of sewage biosolids loaded with SARS-CoV-2 have potential health risks.

Quantitative microbial risk assessment (QMRA) of SARS-CoV-2 in sewage was conducted at the entrance of a WWTP with viral load of 1.03 x 10 2 to 1.31 x 10 4 genome copies/mL (Zaneti et al., 2020) . QMRA was performed to estimate the risk of infection for workers in a three-tiered approach (moderate, aggressive and extreme COVID-19 spread scenarios). The estimated risk values for aggressive and extreme scenarios are 6.5 x 10 -3 and 3.1 x 10 -2 respectively (Zaneti et al., 2020) . Obtained QMRA values were higher than WHO benchmark for tolerable viral infection risk (10 -3 ). Thus, sewage systems appear to be a possible route of viral transmission (Zaneti et al., 2020) .

Previous reports indicates viral persistence in its active state and could cause severe safety risk to the farmers indulged in irrigation process and it can also affect public J o u r n a l P r e -p r o o f health (Arora et al., 2020; Rosa et al., 2020b) . Thus, validation of the absence SARS-CoV-2 genome in treated effluents becomes necessary (Arora et al., 2020) .

The low number of wastewater treatment facilities raises serious problems for developing countries (Arora et al., 2020) . Under such circumstances, validation of the SARS-CoV-2 genome presence in both treated and untreated wastewaters becomes essential. Thus, both untreated and treated wastewaters with SARS-CoV-2 load can cause large scale viral spread in developing countries, where limited medical facilities are available (Arora et al., 2020) . Places where wastewater is untreated and drinking water purification plants do not operate are particular loci for transmission,

Tertiary and advanced treatment technologies feature chlorination, ozonation, photocatalysis, advanced oxidation processes, filtrations (including reverse osmosis, ultra-, micro-, nano-filtrations) and adsorption for water and wastewater treatment (Patel et al., 2019) . Some of these technologies have also been evaluated for coronaviruses including SARS-CoV-2 removal from aqueous systems (Collivignarelli et al., 2020; Wang et al., 2005) . Disinfection agents including bleach, benzyl alkyl ammonium chloride, chloroxylenol, ethanol, povidine-iodine, hypochlorites, quaternary ammonium salts, hydrogen peroxide, peracetic acids, mono persulfates, and chlorine dioxide were suggested for SARS-CoV-2 disinfection (Chin et al., 2020; USEPA, 2020) . They add cost to wastewater treatment.

Chlorine is one of the earliest and most used disinfection agents for wastewater treatment due to its powerful oxidizing nature (Ghernaout et al., 2018; Ghernaout et al., 2011; Yu-mei et al., 2010) . Common chlorine-based disinfectants are chlorine (liquid), J o u r n a l P r e -p r o o f chlorine dioxide and sodium hypochloride (Wang et al., 2020c) . Chlorine-based disinfectants destroy anabolic pathways of proteins and further neutralize microorganisms including viruses, bacteria, spores, fungi, and Clostridium botulinum. In comparison to chlorine, the chlorine dioxide is 5 times more soluble and has 2.63 times more oxidizing capability (Wang et al., 2020c) . Coronaviruses are highly sensitive to chlorine and seem unstable in presence of chlorine (Rosa et al., 2020b) . SARS coronaviruses have a greater vulnerability to residual chlorine than E. coli and f2 phage (Wang et al., 2020c) . Complete inactivation of SARS coronaviruses can be achieved with residual chlorine > 0.5 mg/L or chlorine dioxide > 2.19 mg/L in 30 minutes (Wang et al., 2005) . SARS-CoV-2 inactivation using diluted household bleach (1:99) was performed in vitro and complete inactivation was achieved in 5 minutes contact time (Chin et al., 2020) . 100 % SARS-CoV-2 removal was also achieved through tertiary wastewater treatment equipped with both NaClO, and NaClO coupled with UV irradiation (Randazzo et al., 2020b) . These studies provide insights into the potential of chlorination based process to successfully disinfect aqueous sources contaminated with SARS-CoV-2. However, present research does not clarify facts about required dosage and contact time for viral disinfection in most aqueous systems (Collivignarelli et al., 2020) .

Disinfection agents including ethanol (78-95 %), 2-propanol (70-100 %), 2propanolol and 1-propanol in combination (45 % + 30 %), formaldehyde (0.7-1 %), glutardialdehyde (0.5-2.5 %) and povidone iodine (0.23-7.5 %) can rapidly inactivate the infectivity of coronaviruses by >4 log 10 in suspensions (Kampf et al., 2020a) . Sodium hypochlorite (>0.2 %), hydrogen peroxide (0.5 %) can also disinfect coronavirus J o u r n a l P r e -p r o o f contaminated water (Kampf et al., 2020a) . These doses were based on previous studies used for inactivation of different coronaviruses strains including SARS-CoV-1, HCoV and MERS (Kampf et al., 2020a) . However, detailed studies are still lacking for SARS-CoV-2 inactivation. One study from the Wuchang Cabin hospital in China provides striking revelations about challenges of hospital wastewater disinfection (Zhang et al., 2020a) . Unexpectedly high (0.5-18.7)×10 3 SARS-CoV-2 genome copies/L were found even after disinfection with an 800 g/m 3 sodium hypochlorite dose, which is recommended by WHO and China CDC (Zhang et al., 2020a) . Complete viral inactivation is only achieved with a dose of 6700 g/m 3 sodium hypochlorite dose application (Zhang et al., 2020a) .

Several studies have been performed to inactivate SARS-CoV-1 in presence of high temperatures (Darnell et al., 2004; Darnell and Taylor, 2006; Kariwa et al., 2006; Scheller et al., 2020) . When treated at 56 °C for different time periods, SARS-CoV-1 rate of infection was below the limit of detection (LOD) after a 20 min incubation time.

However, complete inactivation is only possible after 60 min or greater incubation periods (Scheller et al., 2020) . Complete SARS-CoV inactivation was also reported at 75 °C in 45 min (Scheller et al., 2020) . Total inactivation of the virus (>6 Log 10 decrease) was observed at 92 °C after 15 min (Pastorino et al., 2020) . Similar trends were also observed in previous studies on SARS-CoV-1 and MERS-CoV (Darnell et al., 2004; Leclercq et al., 2014) . The effect of a wide temperature range (37, 42, 56, and 60 °C) on SARS-CoV-2 inactivation is reported in SARS-CoV-2 inactivation studies using heat in water and wastewater were almost completely absent. However, wastewater monitoring studies have successfully applied heat for SARS-CoV-2 inactivation for the safety of scientific personal (Arora et al., 2020; Rosa et al., 2020b) . For example, thermal inactivation were performed by treating wastewater samples at 56 °C for 30 minutes (Rosa et al., 2020b) . Other studies inactivated SARS-CoV-2 in wastewater samples using heat at 60 °C for 90 minutes (Arora et al., 2020; Wu et al., 2020a) . (Wang et al., 2020c) . UV wavelength between 200 and 300 nm can destroy the structure of RNA and DNA of viruses, bacteria, and single-celled microorganisms. Therefore, the synthesis of protein is blocked (Ghernaout and Elboughdiri, 2020a; Ghernaout and Elboughdiri, 2020b) . A wavelength of ~254 nm is considered optimum for microbial disinfection through UV J o u r n a l P r e -p r o o f (Meulemans, 1987) . In comparison to chlorine disinfection, UV disinfection requires considerably smaller operation costs and investments . Using UV-C has disadvantages of low depth penetration and possible personal health risks . However, large scale applications have been successfully demonstrated using UV for wastewater disinfections (Hassen et al., 2000) . (According to a study, SARS-CoV-1 viral stocks placed in 24-well tissue culture plates (depth = 1 cm) suffers no significant effects of UV-A (365 nm emitting 2133 μW/cm 2 at a distance of 3 cm) on its infectivity even after 15 min exposure. By contrast, UV-C (254 nm emitting 4016 μW/cm 2 at a distance of 3 cm) partially inactivated the virus in 1 min and provided complete viral inactivation in 15 min (Darnell et al., 2004) .

Gamma radiation has been used for as an effective pathogen inactivation method for decades (Grieb et al., 2002) . Gamma radiation acts by two mechanisms: (i) direct energy transfer by photons of the irradiations, (ii) inactivation of biological material via dislocation of electrons, covalent bond breakage, or by free radicals causing indirect damage (Grieb et al., 2002) . Gamma radiation was also evaluated for the inactivation of SARS-CoV-1 (Darnell et al., 2004) . A SARS-CoV-1 solution (400 μL of 10 6.33 TCID 50 per mL) was subjected to 30, 50, 100, and 150 Gray (1 Gy = J/Kg) of gamma radiation (source 60 Co). But, viral infectivity did not have any significant effect even after 15 min exposure of gamma radiations (Darnell et al., 2004) . A recent review of different solar energy systems for viral disinfection included 1) direct UV, and heat exposure. 2) photocatalytic/thermocatalytic/combined methods for disinfection for use in SARS-CoV-2 inactivation in water (Chauhan, 2020) . Use of plasma discharge has also being suggested for SARS-CoV-2 inactivation (Ghernaout and Elboughdiri, 2020a) .

J o u r n a l P r e -p r o o f

It has been 100 years since the Spanish Flu (H1N1 influenza A) virus swept around the world. In 1918, recommended defensive actions included quarantines, social distancing, banning of mass gatherings and the wearing of face masks (Tomes, 2010) .

There were no antiviral or antibiotics to treat the Spanish Flu virus or consequential infections. Today"s world is more scientifically advanced, and scientists are focused on understanding, managing, treating, and eradicating the new COVID-19 disease. With the might of the modern scientific world mobilized, the amount of data being produced is massive and more is being added daily.

Herein we provided a literature snapshot focused on the occurrence, persistence, analysis, and possible management of coronavirus (SARS-CoV-2) in the aquatic systems. The reader will understand where the virus can be found in the environment including waters, wastewaters, and sewage sludge. We describe the range of methods used for virus sample collection, preservation, preparation, extraction, analysis, and detection. Virus eradication strategies are also addressed for wastewater treatment plants and other aquatic systems using advanced disinfection strategies, heat, and radiation. The questions and recommendations that follow are based on perceived research gaps in the exiting literature. 7. The emergence of wastewater analysis studies in many countries to monitor SARS-CoV-2 is reported in Table 2 . Still, proper WBE studies are lacking at both regional and global level to monitor pandemic situations. Several agencies from Australia, USA and European countries including Netherlands, Britain and Spain are formalizing/started national initiatives to develop early warning systems to tackle future COVID-like outbreaks. It"s been more than 6 months since, the pandemic has spread over the entire planet and no proper WBE system has been developed and implemented to tackle situations like COVID. National and regional health agencies must develop WBE systems to monitor and warn citizens of possible hotspots and rising pandemic situations. These studies must be extended to archived samples as well. where were all branches of the omnipotent news media hiding on this topic?

Sequential concentration of bacteria and viruses from marine waters using a dual membrane system

Stability of SARS-CoV2 and other coronaviruses in the environment and on common touch surfaces

Viromic analysis of wastewater input to a river catchment reveals a diverse assemblage of RNA viruses

First confirmed detection of SARS-CoV-2 in untreated wastewater in Australia: A proof of concept for the wastewater surveillance of COVID-19 in the community

Comparison of virus concentration methods for the RT-qPCR-based recovery of murine hepatitis virus, a surrogate for SARS-CoV-2 from untreated wastewater

SARS-CoV-2 Detection in Sewage in Santiago, Chile-Preliminary results

Sewage surveillance for the presence of SARS-CoV-2 genome as a useful wastewater based epidemiology (WBE) tracking tool in India

The fate of SARS-CoV-2 in wastewater treatment plants points out the sludge line as a suitable spot for incidence monitoring

Identification of viral pathogen diversity in sewage sludge by metagenome analysis

Survival and transport of enteric viruses in the environment. Viruses in foods

Recreational waters-A potential transmission route for SARS-CoV-2 to humans?

Indirect virus transmission in cluster of COVID-19 cases

A Case Series of children with 2019 novel coronavirus infection: clinical and epidemiological features

Persistence of SARS-CoV-2 in the environment and COVID-19 transmission risk from environmental matrices and surfaces

Survival of surrogate coronaviruses in water

Effects of air temperature and relative humidity on coronavirus survival on surfaces

Improved molecular diagnosis of COVID-19 by the novel, highly sensitive and specific

RdRp/Hel real-time reverse transcription-PCR assay validated in vitro and with clinical specimens

A familial cluster of pneumonia associated with the 2019 novel coronavirus indicating person-to-person transmission: a study of a family cluster

Can Solar Energy take part in fighting against COVID-19 pandemic? A review on inactivation of SARS-CoV-2 in Water and Air using Solar Energy

Sentinel surveillance of SARS-CoV-2 in wastewater anticipates the occurrence of COVID-19 cases

Stability of SARS-CoV-2 in different environmental conditions

SARS-CoV-2 in wastewater treatment plants

Viral shedding and antibody response in 37 patients with Middle East respiratory syndrome coronavirus infection

The COVID-19 pandemic: considerations for the waste and wastewater services sector

Inactivation of the coronavirus that induces severe acute respiratory syndrome, SARS-CoV

Evaluation of inactivation methods for severe acute respiratory syndrome coronavirus in noncellular blood products

The international imperative to rapidly and inexpensively monitor community-wide Covid-19 infection status and trends

Wastewater surveillance for population-wide Covid-19: The present and future

SARS-CoV-2 in environmental samples of quarantined households

Aerosol and surface stability of SARS-CoV-2 as compared with SARS-CoV-1

Emerging technologies for the rapid detection of enteric viruses in the aquatic environment

Wastewater and public health: the potential of wastewater surveillance for monitoring COVID-19

Comparative dynamic aerosol efficiencies of three emergent coronaviruses and the unusual persistence of SARS-CoV-2 in aerosol suspensions

Quantifying SARS-CoV-2 transmission suggests epidemic control with digital contact tracing

SARS-CoV-2 from faeces to wastewater treatment: What do we know? A review

SARS-CoV-2 in human sewage in Santa Catalina, Brazil

Production, use, and fate of all plastics ever made

The lethal water tri-therapy: Chlorine, alum, and polyelectrolyte

Disinfecting Water: Plasma Discharge for Removing Coronaviruses

Urgent Proposals for Disinfecting Hospital Wastewaters during COVID-19 Pandemic

On the dependence of chlorine by-products generated species formation of the electrode material and applied charge during electrochemical water treatment

Exaggerated risk of transmission of COVID-19 by fomites

COVID-19 Surveillance in Southeastern Virginia Using Wastewater-Based Epidemiology

CoV-2: The Growing Case for Potential Transmission in a Building via Wastewater Plumbing Systems

COVID-19: mitigating transmission via wastewater plumbing systems

An assessment of, and response to, potential cross-contamination routes due to defective appliance water trap seals in building drainage systems

Environmental conditions and the prevalence of norovirus in hospital building drainage system wastewater and airflows

Quantification of SARS-CoV-2 and cross-assembly phage (crAssphage) from wastewater to monitor coronavirus transmission within communities

Effective use of gamma irradiation for pathogen inactivation of monoclonal antibody preparations

Rios-Touma B. First SARS-CoV-2 detection in river water: implications in low sanitation countries

Survival of coronaviruses in water and wastewater

Persistent viral shedding of SARS-CoV-2 in faeces-a rapid review

Single-Use Plastics and COVID-19: Scientific Evidence and Environmental Regulations

A review on recent progress in the detection methods and prevalence of human enteric viruses in water

First environmental surveillance for the presence of SARS-CoV-2 RNA in wastewater and river water in Japan

Coronavirus disease 2019 (COVID-19): A literature review

Computational analysis of SARS-CoV-2/COVID-19 surveillance by wastewater-based epidemiology locally and globally: Feasibility, economy, opportunities and challenges

Modeling wastewater temperature and attenuation of sewage-borne biomarkers globally

Simulated 2017 nationwide sampling at 13,940 major US sewage treatment plants to assess seasonal population bias in wastewater-based epidemiology

UV disinfection of treated wastewater in a large-scale pilot plant and inactivation of selected bacteria in a laboratory UV device

Detection of SARS-CoV-2 in wastewater in Japan by multiple molecular assays-implication for wastewater-based epidemiology (WBE)

Potential Sensitivity of Wastewater Monitoring for SARS-CoV-2: Comparison with Norovirus Cases

COVID-19 faecal-oral transmission: Are we asking the right questions?

COVID-19: faecal-oral transmission?

First case of 2019 novel coronavirus in the United States

Viral loads in clinical specimens and SARS manifestations

Microfluidic quantitative PCR for simultaneous quantification of multiple viruses in environmental water samples

Strategic and perspectives to develop SARS-CoV-2 detection methods and diagnostics

Simultaneous detection of five enteric viruses associated with gastroenteritis by use of a PCR assay: a single real-time multiplex reaction and its clinical application

Potential role of inanimate surfaces for the spread of coronaviruses and their inactivation with disinfectant agents

Persistence of coronaviruses on inanimate surfaces and their inactivation with biocidal agents

Inactivation of coronaviruses by heat

Probable evidence of fecal aerosol transmission of SARS-CoV-2 in a high-rise building

Inactivation of SARS coronavirus by means of povidoneiodine, physical conditions and chemical reagents

Stability of SARS-CoV-2 on Critical Personal Protective Equipment

First Data-Set on SARS-CoV-2 Detection for Istanbul Wastewaters in Turkey

SARS-CoV-2 Detection in Istanbul Wastewater Treatment Plant Sludges

The first proof of the capability of wastewater surveillance for COVID-19 in India through the detection of the genetic material of SARS-CoV-2

Emission strength of airborne pathogens during toilet flushing

The incubation period of coronavirus disease 2019 (COVID-19) from publicly reported confirmed cases: estimation and application

Heat inactivation of the Middle East respiratory syndrome coronavirus

Clinical and virological data of the first cases of COVID-19 in Europe: a case series

Enteric involvement of severe acute respiratory syndrome-associated coronavirus infection

Can a toilet promote virus transmission? From a fluid dynamics perspective

Persistence and clearance of viral RNA in 2019 novel coronavirus disease rehabilitation patients

The reproductive number of COVID-19 is higher compared to SARS coronavirus

Possible transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a public bath center in Huai'an

Letter to the Editor Regarding:-An Imperative Need for Research on the Role of Environmental Factors in Transmission of Novel Coronavirus (COVID-19)‖-Secondhand and Thirdhand Smoke As Potential Sources of COVID-19

Intensified environmental surveillance supporting the response to wild poliovirus type 1 silent circulation in Israel

Can a Paper-Based Device Trace COVID-19 Sources with Wastewater-Based Epidemiology?

Presence of SARS-Coronavirus-2 RNA in sewage and correlation with reported COVID-19 prevalence in the early stage of the epidemic in the Netherlands

Droplets and Aerosols in the Transmission of SARS-CoV-2

The basic principles of UV-disinfection of water

Sewage analysis as a tool for the COVID-19 pandemic response and management: the urgent need for optimised protocols for SARS-CoV-2 detection and quantification

Estimating the asymptomatic proportion of coronavirus disease 2019 (COVID-19) cases on board the Diamond Princess cruise ship

Preliminary Study of Sars-Cov-2 Occurrence in Wastewater in the Czech Republic

Editorial Perspectives: 2019 novel coronavirus (SARS-CoV-2): what is its fate in urban water cycle and how can the water research community respond? Environmental Science

Comparative performance of SARS-CoV-2 detection assays using seven different primer-probe sets and one assay kit

Temporal detection and phylogenetic assessment of SARS-CoV-2 in municipal wastewater

Estimation of the asymptomatic ratio of novel coronavirus infections (COVID-19)

What do we know about the SARS-CoV-2 coronavirus in the environment?

Air, surface environmental, and personal protective equipment contamination by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from a symptomatic patient

Regressing SARS-CoV-2 sewage measurements onto COVID-19 burden in the population: a proof-of-concept for quantitative environmental surveillance

Viral load of SARS-CoV-2 in clinical samples

Evaluation of heating and chemical protocols for inactivating SARS-CoV-2

Pharmaceuticals of emerging concern in aquatic systems: chemistry, occurrence, effects, and removal methods

A murine herpesvirus closely related to ubiquitous human herpesviruses causes T-cell depletion

SARS-CoV-2 RNA concentrations in primary municipal sewage sludge as a leading indicator of COVID-19 outbreak dynamics

Clinical progression and viral load in a community outbreak of coronavirus-associated SARS pneumonia: a prospective study

Preliminary results of SARS-CoV-2 detection in sewerage system in Niterói municipality

Metropolitan Wastewater Analysis for COVID-19 Epidemiological Surveillance

SARS-CoV-2 RNA in wastewater anticipated COVID-19 occurrence in a low prevalence area

Presence and vitality of SARS-CoV-2 virus in wastewaters and rivers

Preventing SARS-CoV-2 transmission in rehabilitation pools and therapeutic water environments

Coronavirus in water environments: Occurrence, persistence and concentration methods-A scoping review

Emerging and potentially emerging viruses in water environments

First detection of SARS-CoV-2 in untreated wastewaters in Italy

Transmission of 2019-nCoV infection from an asymptomatic contact in Germany

Association of social distancing, population density, and temperature with the instantaneous reproduction number of SARS-CoV-2 in counties across the United States

Aerosol and surface contamination of SARS-CoV-2 observed in quarantine and isolation care

Physicochemical properties of SARS-CoV-2 for drug targeting, virus inactivation and attenuation, vaccine formulation and quality control

Presence of Live SARS-CoV-2 Virus in Feces of Coronavirus Disease 2019 (COVID-19) Patients: A Rapid Review

SARS-Cov-2RNA Found on Particulate Matter of Bergamo in Northern Italy: First Evidence

Detection of SARS-Coronavirus-2 in wastewater, using the existing environmental surveillance network: An epidemiological gateway to an early warning for COVID-19 in communities

First detection of SARS-CoV-2 RNA in wastewater in North America: A study in Louisiana, USA

Risk of SARS-CoV-2 infection from contaminated water systems

Future perspectives of wastewater-based epidemiology: monitoring infectious disease spread and resistance to the community level

Detection and survival of SARS-coronavirus in human stool, urine, wastewater and sludge

Isolation of infectious SARS-CoV-2 from urine of a COVID-19 patient

Early release-Detection of novel coronavirus By RT-PCR in stool specimen from asymptomatic child

Destroyer and teacher‖: Managing the masses during the 1918-1919 influenza pandemic

Disinfectants for Use Against SARS-CoV-2 (COVID-19)

Analytical sensitivity and efficiency comparisons of SARS-COV-2 qRT-PCR assays

Virus transmission from urinals

Disinfection technology of hospital wastes and wastewater: Suggestions for disinfection strategy during coronavirus Disease 2019 (COVID-19) pandemic in China

Effective Heat Inactivation of SARS-CoV-2. medRxiv

Study on the resistance of severe acute respiratory syndrome-associated coronavirus

Emerging infectious diseases: Focus on infection control issues for novel coronaviruses (Severe Acute Respiratory Syndrome-CoV and Middle East Respiratory Syndrome-CoV), hemorrhagic fever viruses (Lassa and Ebola), and highly pathogenic avian influenza viruses, A (H5N1) and A (H7N9)

Consensus document on the epidemiology of severe acute respiratory syndrome (SARS)

Coronavirus disease (COVID-19) pandemic

WHO. WHO Coronavirus Disease (COVID-19) Dashboard

Environmental engineers and scientists have important roles to play in stemming outbreaks and pandemics caused by enveloped viruses

Virological assessment of hospitalized patients with COVID-2019

SARS-CoV-2 titers in wastewater are higher than expected from clinically confirmed cases

Prolonged presence of SARS-CoV-2 viral RNA in faecal samples

Evaluation of lockdown impact on SARS-CoV-2 dynamics through viral genome quantification in Paris wastewaters

Time course quantitative detection of SARS-CoV-2 in Parisian wastewaters correlates with COVID-19 confirmed cases

Evidence for gastrointestinal infection of SARS-CoV-2

A longitudinal survey for genome-based identification of SARS-CoV-2 in sewage water in selected lockdown areas of Lahore city, Pakistan; a potential approach for future smart lockdown strategy

Survivability, partitioning, and recovery of enveloped viruses in untreated municipal wastewater

Enteric involvement of coronaviruses: is faecal-oral transmission of SARS-CoV-2 possible?

Application progress of hospital wastewater treatment methods

QMRA of SARS-CoV-2 for workers in wastewater treatment plants

Potential spreading risks and disinfection challenges of medical wastewater by the presence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) viral RNA in septic tanks of fangcang hospital

Isolation of 2019-nCoV from a stool specimen of a laboratory-confirmed case of the coronavirus disease 2019 (COVID-19)

Viral load dynamics and disease severity in patients infected with SARS-CoV-2 in Zhejiang province, China

Financial support [DST/TM/WTI/2K15/121 (C) dated: 19.09.2016] in the project entitled "Removal and Recovery of Pharmaceuticals from water using sustainable magnetic and nonmagnetic biochars" from Department of Science and Technology, New Delhi, India is thankfully acknowledged. Authors are also thankful to University Grant Commission (UGC), New Delhi for providing the financial assistance under 21st Century Indo-US Research Initiative 2014 to Jawaharlal Nehru University, New Delhi and Mississippi State University, USA in the project "Clean Energy and Water Initiatives" [UGC No. F.194-1/2014(IC)]. One of the authors (DM) also thanks to Jawaharlal Nehru University for providing financial assistance under the Second phase of a University with Potential of Excellence (UPOEII) grant (ID 189). MP is thankful to CSIR for providing financial assistance under CSIR-SRF. Authors also acknowledge the funding support from DST PURSE, Government of India.